Analysis of the oligosaccharides on androgen-binding proteins: Implications concerning their role in structure/function relationships

Rat androgen-binding protein (rABP), human testosterone-binding globulin (hTeBG) and rabbit (rb) TeBG are heterodimeric proteins. The source of the heterogeneity arises from the differential glycosylation of a common protein core. This glycosylation results in a heavy subunit (more glycosylation) and a light subunit (less glycosylation). Glycosylation is one factor responsible for multiple charged species seen when rABP, hTeBG, and rbTeBG are analyzed by two-dimensional gel electrophoresis. Enzymatic digestion with the endoglycosidase, peptide: N-glycosidase F indicated that all three proteins have asparagine (Asn)-linked oligosaccharides as their major glycan substituent. Treatment with exoglycosidases provided evidence for terminal sialic acid, galactose and mannose and N-acetylglucosamine residues. About 16–22% of the mass of the heavy subunit and about 8–14% of the mass of the light subunit is contributed by carbohydrate.

Serial lectin chromatography indicated that rABP is glycosylated differently from hTeBG and rbTeBG. About 40% of the rABP contains tri and tetraantennary complex oligosaccharides, while only about 20% of the hTeBG and TeBG from pregnant rabbits contains these types of glycans. About 9% of the TeBG from male rabbits bears these types of oligosaccharides. All of the biantennary complex oligosaccharides on rABP are fucosylated on the chitobiose core, but only 8% of those on hTeBG and none of those on rbTeBG are fucosylated in this manner. All three proteins are glycosylated at more than one site. The data indicate that the proteins may have more than one type of oligosaccharide on them. It is likely that differences in glycosylation are responsible for different physiological roles of the proteins.     

A simple,rapid, and sensitive DNA assay procedure

A simple and rapid assay for quantitative determinations of DNA in crude homogenates is described. The method is based on the enhancement of fluorescence seen when bisbenzimidazole (Hoechst 33258) binds to DNA. Crude homogenates in which chromatin has been dissociated with high salt buffer can be assayed directly and reliably in a few minutes. The dissociation of chromatin is critical to accurate determinations of DNA in biological materials using this method. The assay can detect as little as 10 ng of DNA with rather unsophisticated instrumentation.     

Genetic Regulation of Mup Production in Recombinant Inbred Mice

Inbred strains of mice excrete all three major urinary proteins (mups) when induced by testosterone, but differ as to the relative proportions and total levels of each mup present. We have now determined the urinary mup phenotypes before and after testosterone treatment of seven recombinant inbred strains derived from progenitor strains exhibiting different mup phenotypes. The results confirm previous observations indicating that total control of mup protein production is a multigenic process. One locus, Mup-a on chromosome 4, determines the relative mup protein proportions after induction by testosterone. Mup-a, together with other genetic sites, determines the basal mup proportions. Genes other than Mup-a determine the kinetics of mup induction and total mup excretion.     

Observations of a set of isomeric anti-lactose antibodies directed against a group D streptococcal polysaccharide

Sets of isomeric anti-lactose antibodies with specificity for the lactose units of a cell wall polysaccharide fromStreptococcus faecalis strain N were induced in rabbits immunized with a vaccine of nonviable cells of the organism. Such sets of anti-lactose antibodies were isolated from the serum of immunized animals by affinity chromatography on lactosyl-Sepharose. Gel electrofocusing experiments showed that the preparations consisted of multiprotein components. One preparation of antibodies of 13 isomers was separated into homogeneous components by liquid isoelectrofocusing. The individual isomeric antibodies exhibit specificity for the lactose units of the antigenic polysaccharide, possess isoelectric points in the range of 5.9–8.0, and belong to the IgG class of immunoglobulins, and each member yields one light chain and one heavy chain on dissociation in sodium dodecyl sulfate (SDS) and mercaptoethanol. These results have been interpreted as evidence for the assembly of the chains of isomeric antibodies by a single-chain pairing mechanism.     

A Systematic Review of the Relationship between Blood Loss and Clinical Signs

Introduction

This systematic review examines the relationship between blood loss and clinical signs and explores its use to trigger clinical interventions in the management of obstetric haemorrhage.

Methods

A systematic review of the literature was carried out using a comprehensive search strategy to identify studies presenting data on the relationship of clinical signs & symptoms and blood loss. Methodological quality was assessed using the STROBE checklist and the general guidelines of MOOSE.

Results

30 studies were included and five were performed in women with pregnancy-related haemorrhage (other studies were carried in non-obstetric populations). Heart rate (HR), systolic blood pressure (SBP) and shock index were the parameters most frequently studied. An association between blood loss and HR changes was observed in 22 out of 24 studies, and between blood loss and SBP was observed in 17 out of 23 studies. An association was found in all papers reporting on the relationship of shock index and blood loss. Seven studies have used Receiver Operating Characteristic Curves to determine the accuracy of clinical signs in predicting blood loss. In those studies the AUC ranged from 0.56 to 0.74 for HR, from 0.56 to 0.79 for SBP and from 0.77 to 0.84 for shock index. In some studies, HR, SBP and shock index were associated with increased mortality.

Conclusion

We found a substantial variability in the relationship between blood loss and clinical signs, making it difficult to establish specific cut-off points for clinical signs that could be used as triggers for clinical interventions. However, the shock index can be an accurate indicator of compensatory changes in the cardiovascular system due to blood loss. Considering that most of the evidence included in this systematic review is derived from studies in non-obstetric populations, further research on the use of the shock index in obstetric populations is needed.     

Strong and weak plasma response to dietary carotenoids identified by cluster analysis and linked to beta-carotene 15,15'-monooxygenase 1 single nucleotide polymorphisms

The mechanisms as well the genetics underlying the bioavailability and metabolism of carotenoids in humans remain unclear. To begin to address these questions, we used cluster analysis to examine individual temporal responses of plasma carotenoids from a controlled-diet study of subjects who consumed carotenoid-rich beverages. Treatments, given daily for 3 weeks, were watermelon juice at two levels (20-mg lycopene, 2.5-mg β-carotene, n=23 and 40-mg lycopene, 5-mg β-carotene, n=12) and tomato juice (18-mg lycopene, 0.6-mg β-carotene, n=10). Cluster analysis revealed distinct groups of subjects differing in the temporal response of plasma carotenoids and provided the basis for classifying subjects as strong responders or weak responders for β-carotene, lycopene, phytoene and phytofluene. Individuals who were strong or weak responders for one carotenoid were not necessarily strong or weak responders for another carotenoid. Furthermore, individual responsiveness was associated with genetic variants of the carotenoid metabolizing enzyme β-carotene 15,15'-monooxygenase 1. These results support the concept that individuals absorb or metabolize carotenoids differently across time and suggest that bioavailability of carotenoids may involve specific genetic variants of β-carotene 15,15'-monooxygenase 1.